Temozolomide increases the generation of cell heterogeneity in ERK activity in glioma cells.

ERK activity governs diverse cellular responses and has significant implications in cancer biology and treatment. Cellular heterogeneity is a major feature of cancer and a barrier for therapy success, allowing cancer cells to adapt and survive in challenging environments. Here, we used a genetic live-cell reporter to explore the heterogeneity of ERK signaling activity within cellular populations and colonies of glioblastoma (GB) cells. GB cells showed a wide spectrum of ERK activation levels in basal culture conditions and throughout state transitions. Treatment with the chemotherapeutic agent temozolomide increased the phenotypic heterogeneity in ERK activity within cells even in clonal populations. Using the MEK inhibitor trametinib in combination with temozolomide to homogenize ERK activity reduced cell fitness in colonies and decreased fractional killing in GB clonal cells. Our study contributes to the growing understanding of the complexity in ERK activity and dynamics, pointing out the consequences of cell-to-cell ERK phenotypic variability in fitness and therapy survival. The complexity of ERK signaling phenotypes in the context of chemotherapy treatment is shown, offering valuable insights about the intricacies of ERK signaling heterogeneity and chemotherapy treatment. KEY MESSAGES: Heterogeneity in ERK activity in live GBM cells is high in basal culture conditions. Temozolomide alters ERK activity in GBM cells and generates phenotypic heterogeneity. Clonal populations behave heterogeneously in ERK activity, and this heterogeneity impacts fitness. Targeting the generation of ERK heterogeneity reduces fitness and fractional killing in GBM colonies.